Elucidation of molecular pathways that determine renin secretion is of critical significance in understanding the role of renin-angiotensin system in the control of extracellular volume and blood pressure. Macula densa (MD) control of renin secretion is one of the key renin regulating pathways, in in vitro perfused juxtaglomerular apparatus (JGA), a reduction of NaC4 concentration at the MD induced a marked increase in renin secretion. The mediators responsible for the MD control of renin secretion have been recognized to be prostaglandins and nitric oxide. In support of this notion, cyclooxygenase-2 (COX-2) and neuronal form of nitric oxide synthase (nNOS) are co-expressed in the MD, stimulated in parallel in various high renin states, both being implicated to play a role in the control of renin secretion. However, the interaction of the two systems in the control of renin secretion is poorly defined. Our preliminary studies showed that plasma renin activity was significantly reduced in both COX-2 and nNOS knockout mice. In a recently established macula densa cell line, nitric oxide stimulated COX-2 expression and in turn the COX-2 product PGE2 inhibited nNOS expression that was further supported by the finding that nNOS expression was upregulated in the MD of COX-2 knockout mice. We hypothesize that the distinct interaction of COX-2 and nNOS in the MD cells forms a negative feedback loop that may play an important role in the tight control of renin secretion. The major goals of this proposal are to further delineate this feedback loop and to clarify its role in regulation of renin secretion with the following specific aims: 1) investigate the molecular mechanisms by which the COX-2 product PGE2 down-regulates nNOS expression in cultured macula densa cells in vitro; 2) investigate the molecular mechanisms by which the nNOS product nitric oxide stimulates COX-2 expression or activity in the cultured macula densa cells in vitro; and 3) further clarify the interrelationship among COX-2, renin and NOS isoforrns in vivo.